Targeting glutamine dependence with DRP-104 inhibits proliferation and tumor growth of castration-resistant prostate cancer.

BACKGROUND: Prostate cancer (PCa) continues to be one of the leading causes of cancer deaths in men. While androgen deprivation therapy is initially effective, castration-resistant PCa (CRPC) often recurs and has limited treatment options. Our previous study identified glutamine metabolism to be critical for CRPC growth. The glutamine antagonist 6-diazo-5-oxo-l-norleucine (DON) blocks both carbon and nitrogen pathways but has dose-limiting toxicity. The prodrug DRP-104 is expected to be preferentially converted to DON in tumor cells to inhibit glutamine utilization with minimal toxicity. However, CRPC cells' susceptibility to DRP-104 remains unclear. METHODS: Human PCa cell lines (LNCaP, LAPC4, C4-2/MDVR, PC-3, 22RV1, NCI-H660) were treated with DRP-104, and effects on proliferation and cell death were assessed. Unbiased metabolic profiling and isotope tracing evaluated the effects of DRP-104 on glutamine pathways. Efficacy of DRP-104 in vivo was evaluated in a mouse xenograft model of neuroendocrine PCa, NCI-H660. RESULTS: DRP-104 inhibited proliferation and induced apoptosis in CRPC cell lines. Metabolite profiling showed decreases in the tricarboxylic acid cycle and nucleotide synthesis metabolites. Glutamine isotope tracing confirmed the blockade of both carbon pathway and nitrogen pathways. DRP-104 treated CRPC cells were rescued by the addition of nucleosides. DRP-104 inhibited neuroendocrine PCa xenograft growth without detectable toxicity. CONCLUSIONS: The prodrug DRP-104 blocks glutamine carbon and nitrogen utilization, thereby inhibiting CRPC growth and inducing apoptosis. Targeting glutamine metabolism pathways with DRP-104 represents a promising therapeutic strategy for CRPC.

Whole-Slide Cytometric Feature Mapping for Distinguishing Tumor Genomic Subtypes in Head and Neck Squamous Cell Carcinoma Whole-Slide Images.

Head and neck squamous cell carcinoma (HNSCC) is a heterogeneous disease where, in advanced stages, clinical and pathologic stages do not correlate with outcome. Molecular and genomic biomarkers for HNSCC classification have shown promise for prognostic and therapeutic applications. This study utilized automated image analysis techniques in whole-slide images of HNSCC tumors to identify relationships between cytometric features and genomic phenotypes. Hematoxylin and eosin-stained slides of HNSCC tumors (N = 49) were obtained from The Cancer Imaging Archive, along with accompanying clinical, pathologic, genomic, and proteomic reports. Automated nuclear detection was performed across the entirety of slides, and cytometric feature maps were generated. Forty-one cytometric features were evaluated for associations with tumor grade, tumor stage, tumor subsite, and integrated genomic subtype. Thirty-two features demonstrated significant association with integrated genomic subtype when corrected for multiple comparisons. In particular, the basal subtype was visually distinguishable from the chromosomal instability and immune subtypes based on cytometric feature measurements. No features were significantly associated with tumor grade, stage, or subsite. This study provides preliminary evidence that features derived from tissue pathology slides could provide insights into genomic phenotypes of HNSCC.

Cas13d knockdown of lung protease Ctsl prevents and treats SARS-CoV-2 infection.

SARS-CoV-2 entry into cells requires specific host proteases; however, no successful in vivo applications of host protease inhibitors have yet been reported for treatment of SARS-CoV-2 pathogenesis. Here we describe a chemically engineered nanosystem encapsulating CRISPR-Cas13d, developed to specifically target lung protease cathepsin L (Ctsl) messenger RNA to block SARS-CoV-2 infection in mice. We show that this nanosystem decreases lung Ctsl expression in normal mice efficiently, specifically and safely. We further show that this approach extends survival of mice lethally infected with SARS-CoV-2, correlating with decreased lung virus burden, reduced expression of proinflammatory cytokines/chemokines and diminished severity of pulmonary interstitial inflammation. Postinfection treatment by this nanosystem dramatically lowers the lung virus burden and alleviates virus-induced pathological changes. Our results indicate that targeting lung protease mRNA by Cas13d nanosystem represents a unique strategy for controlling SARS-CoV-2 infection and demonstrate that CRISPR can be used as a potential treatment for SARS-CoV-2 infection.

Automated Nuclear Segmentation in Head and Neck Squamous Cell Carcinoma Pathology Reveals Relationships between Cytometric Features and ESTIMATE Stromal and Immune Scores.

The tumor microenvironment (TME) plays an important role in the progression of head and neck squamous cell carcinoma (HNSCC). Currently, pathologic assessment of TME is nonstandardized and subject to observer bias. Genome-wide transcriptomic approaches to understanding the TME, while less subject to bias, are expensive and not currently a part of the standard of care for HNSCC. To identify pathology-based biomarkers that correlate with genomic and transcriptomic signatures of TME in HNSCC, cytometric feature maps were generated in a publicly available data set from a cohort of patients with HNSCC, including whole-slide tissue images and genomic and transcriptomic phenotyping (N = 49). Cytometric feature maps were generated based on whole-slide nuclear detection, using a deep-learning algorithm trained for StarDist nuclear segmentation. Cytometric features in each patient were compared to transcriptomic measurements, including Estimation of Stromal and Immune Cells in Malignant Tumor Tissues Using Expression Data (ESTIMATE) scores and stemness scores. With correction for multiple comparisons, one feature (nuclear circularity) demonstrated a significant linear correlation with ESTIMATE stromal score. Two features (nuclear maximum and minimum diameter) correlated significantly with ESTIMATE immune score. Three features (nuclear solidity, nuclear minimum diameter, and nuclear circularity) correlated significantly with transcriptomic stemness score. This study provides preliminary evidence that observer-independent, automated tissue-slide analysis can provide insights into the HNSCC TME which correlate with genomic and transcriptomic assessments.

KeyLoop retractor for global gasless laparoscopy: evaluation of safety and feasibility in a porcine model.

BACKGROUND: Many surgeons in low- and middle-income countries have described performing surgery using gasless (lift) laparoscopy due to inaccessibility of carbon dioxide and reliable electricity, but the safety and feasibility of the technique has not been well documented. We describe preclinical testing of the in vivo safety and utility of KeyLoop, a laparoscopic retractor system to enable gasless laparoscopy. METHODS: Experienced laparoscopic surgeons completed a series of four laparoscopic tasks in a porcine model: laparoscopic exposure, small bowel resection, intracorporeal suturing with knot tying, and cholecystectomy. For each participating surgeon, the four tasks were completed in a practice animal using KeyLoop. Surgeons then completed these tasks using standard-of-care (SOC) gas laparoscopy and KeyLoop in block randomized order to minimize learning curve effect. Vital signs, task completion time, blood loss and surgical complications were compared between SOC and KeyLoop using paired nonparametric tests. Surgeons completed a survey on use of KeyLoop compared to gas laparoscopy. Abdominal wall tissue was evaluated for injury by a blinded pathologist. RESULTS: Five surgeons performed 60 tasks in 15 pigs. There were no significant differences in times to complete the tasks between KeyLoop and SOC. For all tasks, there was a learning curve with task completion times related to learning the porcine model. There were no significant differences in blood loss, vital signs or surgical complications between KeyLoop and SOC. Eleven surgeons from the United States and Singapore felt that KeyLoop could be used to safely perform several common surgical procedures. No abdominal wall tissue injury was observed for either KeyLoop or SOC. CONCLUSIONS: Procedure times, blood loss, abdominal wall tissue injury and surgical complications were similar between KeyLoop and SOC gas laparoscopy for basic surgical procedures. This data supports KeyLoop as a useful tool to increase access to laparoscopy in low- and middle-income countries.

ß-Cyclodextrin-containing polymer treatment of cutaneous lupus and influenza improves outcomes.

Nucleic acid (NA)-containing damage- and pathogen-associated molecular patterns (DAMPs and PAMPs, respectively) are implicated in numerous pathological conditions from infectious diseases to autoimmune disorders. Nucleic acid-binding polymers, including polyamidoamine (PAMAM) dendrimers, have demonstrated anti-inflammatory properties when administered to neutralize DAMPs/PAMPs. The PAMAM G3 variant has been shown to have beneficial effects in a cutaneous lupus erythematosus (CLE) murine model and improve survival of mice challenged with influenza. Unfortunately, the narrow therapeutic window of cationic PAMAM dendrimers makes their clinical development challenging. An alternative nucleic acid-binding polymer that has been evaluated in humans is a linear ß-cyclodextrin-containing polymer (CDP). CDP's characteristics prompted us to evaluate its anti-inflammatory potential in CLE autoimmune and influenza infectious disease mouse models. We report that CDP effectively inhibits NA-containing DAMP-mediated activation of Toll-like receptors (TLRs) in cell culture, improves healing in lupus mice, and does not immunocompromise treated animals upon influenza infection but improves survival even when administered 3 days after infection. Finally, as anticipated, we observe limited toxicity in animals treated with CDP compared with PAMAM G3. Thus, CDP is a new anti-inflammatory agent that may be readily translated to the clinic to combat diseases associated with pathological NA-containing DAMPs/PAMPs.

Update on the Features and Measurements of Experimental Acute Lung Injury in Animals: An Official American Thoracic Society Workshop Report.

Advancements in methods, technology, and our understanding of the pathobiology of lung injury have created the need to update the definition of experimental acute lung injury (ALI). We queried 50 participants with expertise in ALI and acute respiratory distress syndrome using a Delphi method composed of a series of electronic surveys and a virtual workshop. We propose that ALI presents as a "multidimensional entity" characterized by four "domains" that reflect the key pathophysiologic features and underlying biology of human acute respiratory distress syndrome. These domains are 1) histological evidence of tissue injury, 2) alteration of the alveolar-capillary barrier, 3) presence of an inflammatory response, and 4) physiologic dysfunction. For each domain, we present "relevant measurements," defined as those proposed by at least 30% of respondents. We propose that experimental ALI encompasses a continuum of models ranging from those focusing on gaining specific mechanistic insights to those primarily concerned with preclinical testing of novel therapeutics or interventions. We suggest that mechanistic studies may justifiably focus on a single domain of lung injury, but models must document alterations of at least three of the four domains to qualify as "experimental ALI." Finally, we propose that a time criterion defining "acute" in ALI remains relevant, but the actual time may vary based on the specific model and the aspect of injury being modeled. The continuum concept of ALI increases the flexibility and applicability of the definition to multiple models while increasing the likelihood of translating preclinical findings to critically ill patients.

Safety evaluation of carbon tetrafluoride as an inert hyperbaric breathing gas in Sprague-Dawley rats.

Carbon tetrafluoride (CF4) is an inert gas with higher molecular weight and lower water solubility than commonly used hyperbaric breathing gases. These inert gas properties decrease time required to decompress and avoid decompression sickness after deep dives. To assess CF4 toxicity, Sprague-Dawley rats were exposed to 8 atm absolute (ATA) air (10 males, 10 females) or 8 ATA 79% CF4/21% O2 (25 males, 25 females). Exposures were 30 min daily for 5 days. Rat behavior was normal throughout the testing period. There were no gross or microscopic pathology abnormalities following repeat dose exposure. Male body weight trends were similar between groups. Female body weight trends were 0.5 ± 0.8% day-1 for hyperbaric air exposure and - 0.2 ± 0.8% day-1 for hyperbaric CF4 exposure (P = 0.01) but remained within literature cited norms. Organ weights and hematologic indices remained within or near literature normal ranges. Clinical chemistry panels showed no signs of toxicity in renal or hepatic biomarkers. Polychromatic erythrocyte micronucleus frequency showed no chromosomal damage. Comet assay showed no DNA damage in lung tissue. Females exposed to CF4 had 2.5 times greater percent tail DNA in liver tissue than controls (P = 0.009). However this result remained within the normal range of local negative controls. A bacterial reverse mutation assay with exposure to 1 ATA 79% CF4/21% O2 for 72 h was nonmutagenic in four strains of Salmonella typhimurium and one strain of Escherichia coli. Overall, there was no evidence that CF4 caused organ toxicity or genetic toxicity.

Selected Resources for Pathology Evaluation of Nonhuman Primates in Nonclinical Safety Assessment.

Humans and nonhuman primates (NHPs) share numerous anatomical and physiological characteristics, thereby explaining the importance of NHPs as essential animal models for translational medicine and nonclinical toxicity testing. Researchers, toxicologic pathologists, toxicologists, and regulatory reviewers must be familiar with normal and abnormal NHP biological traits when designing, performing, and interpreting data sets from NHP studies. The current compilation presents a list of essential books, journal articles, and websites that provide context to safety assessment and research scientists working with NHP models. The resources used most frequently by the authors have been briefly annotated to permit readers to rapidly ascertain their applicability to particular research endeavors. The references are aimed primarily for toxicologic pathologists working with cynomolgus and rhesus macaques and common marmosets in efficacy and safety assessment studies.

Proceedings of the 2022 National Toxicology Program Satellite Symposium.

The 2022 annual National Toxicology Program Satellite Symposium, entitled "Pathology Potpourri," was held in Austin, Texas at the Society of Toxicologic Pathology's 40th annual meeting during a half-day session on Sunday, June 19. The goal of this symposium was to present and discuss challenging diagnostic pathology and/or nomenclature issues. This article presents summaries of the speakers' talks along with select images that were used by the audience for voting and discussion. Various lesions and topics covered during the symposium included induced and spontaneous neoplastic and nonneoplastic lesions in the mouse lung, spontaneous lesions in the reproductive tract of a female cynomolgus macaque, induced vascular lesions in a mouse asthma model and interesting case studies in a rhesus macaque, dog and genetically engineered mouse model.

Poly(lactide-co-ε-caprolactone) scaffold promotes equivalent tissue integration and supports skin grafts compared to a predicate collagen scaffold.

Dermal scarring from motor vehicle accidents, severe burns, military blasts, etc. is a major problem affecting over 80 million people worldwide annually, many of whom suffer from debilitating hypertrophic scar contractures. These stiff, shrunken scars limit mobility, impact quality of life, and cost millions of dollars each year in surgical treatment and physical therapy. Current tissue engineered scaffolds have mechanical properties akin to unwounded skin, but these collagen-based scaffolds rapidly degrade over 2 months, premature to dampen contracture occurring 6-12 months after injury. This study demonstrates a tissue engineered scaffold can be manufactured from a slow-degrading viscoelastic copolymer, poly(ι-lactide-co-ε-caprolactone), with physical and mechanical characteristics to promote tissue ingrowth and support skin-grafts. Copolymers were synthesized via ring-opening polymerization. Solvent casting/particulate leaching was used to manufacture 3D porous scaffolds by mixing copolymers with particles in an organic solvent followed by casting into molds and subsequent particle leaching with water. Scaffolds characterized through SEM, micro-CT, and tensile testing confirmed the required thickness, pore size, porosity, modulus, and strength for promoting skin-graft bioincorporation and dampening fibrosis in vivo. Scaffolds were Oxygen Plasma Treatment and collagen coated to encourage cellular proliferation. Porosity ranging from 70% to 90% was investigated in a subcutaneous murine model and found to have no clinical effect on tissue ingrowth. A swine full-thickness skin wound model confirmed through histology and Computer Planimetry that scaffolds promote skin-graft survival, with or without collagen coating, with equal safety and efficacy as a commercially available tissue engineered scaffold. This study validates a scalable method to create poly(ι-lactide-co-ε-caprolactone) scaffolds with appropriate characteristics and confirms in mouse and swine wound models that the scaffolds are safe and effective at supporting skin-grafts. The results of this study have brought us closer towards developing an alternative technology that supports skin grafts with the potential to investigate long-term hypertrophic scar contractures.

Long-term complications of glycogen storage disease type Ia in the canine model treated with gene replacement therapy.

BACKGROUND: Glycogen storage disease type Ia (GSD Ia) in dogs closely resembles human GSD Ia. Untreated patients with GSD Ia develop complications associated with glucose-6-phosphatase (G6Pase) deficiency. Survival of human patients on intensive nutritional management has improved; however, long-term complications persist including renal failure, nephrolithiasis, hepatocellular adenomas (HCA), and a high risk for hepatocellular carcinoma (HCC). Affected dogs fail to thrive with dietary therapy alone. Treatment with gene replacement therapy using adeno-associated viral vectors (AAV) expressing G6Pase has greatly prolonged life and prevented hypoglycemia in affected dogs. However, long-term complications have not been described to date. METHODS: Five GSD Ia-affected dogs treated with AAV-G6Pase were evaluated. Dogs were euthanized due to reaching humane endpoints related to liver and/or kidney involvement, at 4 to 8 years of life. Necropsies were performed and tissues were analyzed. RESULTS: Four dogs had liver tumors consistent with HCA and HCC. Three dogs developed renal failure, but all dogs exhibited progressive kidney disease histologically. Urolithiasis was detected in two dogs; uroliths were composed of calcium oxalate and calcium phosphate. One affected and one carrier dog had polycystic ovarian disease. Bone mineral density was not significantly affected. CONCLUSIONS: Here, we show that the canine GSD Ia model demonstrates similar long-term complications as GSD Ia patients in spite of gene replacement therapy. Further development of gene therapy is needed to develop a more effective treatment to prevent long-term complications of GSD Ia.

HIV-1 Envelope Mimicry of Host Enzyme Kynureninase Does Not Disrupt Tryptophan Metabolism.

The HIV-1 envelope protein (Env) has evolved to subvert the host immune system, hindering viral control by the host. The tryptophan metabolic enzyme kynureninase (KYNU) is mimicked by a portion of the HIV Env gp41 membrane proximal region (MPER) and is cross-reactive with the HIV broadly neutralizing Ab (bnAb) 2F5. Molecular mimicry of host proteins by pathogens can lead to autoimmune disease. In this article, we demonstrate that neither the 2F5 bnAb nor HIV MPER-KYNU cross-reactive Abs elicited by immunization with an MPER peptide-liposome vaccine in 2F5 bnAb VHDJH and VLJL knock-in mice and rhesus macaques modified KYNU activity or disrupted tissue tryptophan metabolism. Thus, molecular mimicry by HIV-1 Env that promotes the evasion of host anti-HIV-1 Ab responses can be directed toward nonfunctional host protein epitopes that do not impair host protein function. Therefore, the 2F5 HIV Env gp41 region is a key and safe target for HIV-1 vaccine development.

Recommendations for minimum information for publication of experimental pathology data: MINPEPA guidelines.

Animal models are essential research tools in modern biomedical research, but there are concerns about their lack of reproducibility and the failure of animal data to translate into advances in human medical therapy. A major factor in improving experimental reproducibility is thorough communication of research methodologies. The recently published ARRIVE guidelines outline basic information that should be provided when reporting animal studies. This paper builds on ARRIVE by providing the minimum information needed in reports to allow proper assessment of pathology data gathered from animal tissues. This guidance covers aspects of experimental design, technical procedures, data gathering, analysis, and presentation that are potential sources of variation when creating morphological, immunohistochemical (IHC) or in situ hybridization (ISH) datasets. This reporting framework will maximize the likelihood that pathology data derived from animal experiments can be reproduced by ensuring that sufficient information is available to allow for replication of the methods and facilitate inter-study comparison by identifying potential interpretative confounders.

The future of preclinical animal models in pharmaceutical discovery and development: a need to bring in cerebro to the in vivo discussions.

Animal models have provided an important tool to help make the decision to take potential therapies from preclinical studies to humans. In the past several years, the strong reliance of the pharmaceutical discovery and development process on the use of animal models has come under increasing scrutiny for ethical and scientific reasons. Several prominent and widely publicized articles have reported limited concordance of animal experiments with subsequent human clinical trials. Recent assessments of the quality of animal studies have suggested that this translational failure may be due in part to shortcomings in the planning, conduct, and reporting of in vivo studies. This article will emphasize methods to assure best practice rigor in animal study methods and reporting. It will introduce the so-called scientific 3Rs of relevance, robustness, and reproducibility to the in vivo study approach and will review important new trends in the animal research and pharmaceutical discovery and development communities.

Pathologists' perspective on the study design, analysis, and interpretation of proliferative lesions in a lifetime rodent carcinogenicity bioassay of sucralose.

Sucralose, a sugar substitute first approved for use in 1991, is a non-caloric sweetener regulated globally as a food additive. Based on numerous experimental animal studies (dating to the 1980s) and human epidemiology studies, international health agencies have determined that sucralose is safe when consumed as intended. A single lifetime rodent carcinogenicity bioassay conducted by the Ramazzini Institute (RI) reported that mice fed diets containing sucralose develop hematopoietic neoplasia, but controversy continues regarding the validity and relevance of these data for predicting health effects in humans. The present paper addresses the controversy by providing the perspective of experienced pathologists on sucralose-related animal toxicity and carcinogenicity data generally, and the RI carcinogenicity bioassay findings specifically, using results from publicly available papers and international regulatory authority decisions. In the authors' view, flaws in the design, methodology, data evaluation, and reporting of the RI carcinogenicity bioassay for sucralose diminish the value of the data as evidence that this agent represents a carcinogenic hazard to humans. This limitation will remain until the RI bioassay is repeated under Good Laboratory Practices and the design, data, and accuracy of the pathology diagnoses and interpretations are reviewed by qualified pathologists with experience in evaluating potential chemically-induced carcinogenic hazards.

Irgm proteins attenuate inflammatory disease in mouse models of genital Chlamydia infection.

Chlamydiae are obligate intracellular bacterial pathogens that may cause genital pathology via induction of destructive host immune responses. Human-adapted Chlamydia trachomatis causes inflammatory disease in human hosts but is easily cleared in mice, and mouse-adapted Chlamydia muridarum establishes a productive and pathogenic infection in murine hosts. While numerous anti-chlamydial host resistance factors have been discovered in mice and humans alike, little is known about host factors promoting host fitness independent of host resistance. Here, we show that interferon-inducible immunity-related GTPase M (Irgm) proteins function as such host factors ameliorating infection-associated sequalae in the murine female genital tract, thus characterizing Irgm proteins as mediators of disease tolerance. Specifically, we demonstrate that mice deficient for all three murine Irgm paralogs (pan-Irgm-/-) are defective for cell-autonomous immunity to C. trachomatis, which correlates with an early and transient increase in bacterial burden and sustained hyperinflammation in vivo. In contrast, upon infection of pan-Irgm-/- mice with C. muridarum, bacterial burden is unaffected, yet genital inflammation and scarring pathology are nonetheless increased, demonstrating that Irgm proteins can promote host fitness without altering bacterial burden. Additionally, pan-Irgm-/- mice display increased granulomatous inflammation in genital Chlamydia infection, implicating Irgm proteins in the regulation of granuloma formation and maintenance. These findings demonstrate that Irgm proteins regulate pathogenic immune responses to Chlamydia infection in vivo, establishing an effective infection model to examine the immunoregulatory functions and mechanisms of Irgm proteins. IMPORTANCE: In response to genital Chlamydia infection, the immune system mounts a proinflammatory response to resist the pathogen, yet inflammation must be tightly controlled to avoid collateral damage and scarring to host genital tissue. Variation in the human IRGM gene is associated with susceptibility to autoinflammatory diseases but its role in ameliorating inflammatory diseases caused by infections is poorly defined. Here, we use mice deficient for all three murine Irgm paralogs to demonstrate that Irgm proteins not only provide host resistance to Chlamydia infections but also limit associated inflammation in the female genital tract. In particular, we find that murine Irgm expression prevents granulomatous inflammation, which parallels inflammatory diseases associated with variants in human IRGM. Our findings therefore establish genital Chlamydia infection as a useful model to study the roles for Irgm proteins in both promoting protective immunity and limiting pathogenic inflammation.

Determining the Relationship between Delivery Parameters and Ablation Distribution for Novel Gel Ethanol Percutaneous Therapy in Ex Vivo Swine Liver.

Ethyl cellulose-ethanol (ECE) is emerging as a promising formulation for ablative injections, with more controllable injection distributions than those from traditional liquid ethanol. This study evaluates the influence of salient injection parameters on forces needed for infusion, depot volume, retention, and shape in a large animal model relevant to human applications. Experiments were conducted to investigate how infusion volume (0.5 mL to 2.5 mL), ECE concentration (6% or 12%), needle gauge (22 G or 27 G), and infusion rate (10 mL/h) impacted the force of infusion into air using a load cell. These parameters, with the addition of manual infusion, were investigated to elucidate their influence on depot volume, retention, and shape (aspect ratio), measured using CT imaging, in an ex vivo swine liver model. Force during injection increased significantly for 12% compared to 6% ECE and for 27 G needles compared to 22 G. Force variability increased with higher ECE concentration and smaller needle diameter. As infusion volume increased, 12% ECE achieved superior depot volume compared to 6% ECE. For all infusion volumes, 12% ECE achieved superior retention compared to 6% ECE. Needle gauge and infusion rate had little influence on the observed depot volume or retention; however, the smaller needles resulted in higher variability in depot shape for 12% ECE. These results help us understand the multivariate nature of injection performance, informing injection protocol designs for ablations using gel ethanol and infusion, with volumes relevant to human applications.

MR histology reveals tissue features beneath heterogeneous MRI signal in genetically engineered mouse models of sarcoma.

Purpose: To identify significant relationships between quantitative cytometric tissue features and quantitative MR (qMRI) intratumorally in preclinical undifferentiated pleomorphic sarcomas (UPS). Materials and methods: In a prospective study of genetically engineered mouse models of UPS, we registered imaging libraries consisting of matched multi-contrast in vivo MRI, three-dimensional (3D) multi-contrast high-resolution ex vivo MR histology (MRH), and two-dimensional (2D) tissue slides. From digitized histology we generated quantitative cytometric feature maps from whole-slide automated nuclear segmentation. We automatically segmented intratumoral regions of distinct qMRI values and measured corresponding cytometric features. Linear regression analysis was performed to compare intratumoral qMRI and tissue cytometric features, and results were corrected for multiple comparisons. Linear correlations between qMRI and cytometric features with p values of <0.05 after correction for multiple comparisons were considered significant. Results: Three features correlated with ex vivo apparent diffusion coefficient (ADC), and no features correlated with in vivo ADC. Six features demonstrated significant linear relationships with ex vivo T2*, and fifteen features correlated significantly with in vivo T2*. In both cases, nuclear Haralick texture features were the most prevalent type of feature correlated with T2*. A small group of nuclear topology features also correlated with one or both T2* contrasts, and positive trends were seen between T2* and nuclear size metrics. Conclusion: Registered multi-parametric imaging datasets can identify quantitative tissue features which contribute to UPS MR signal. T2* may provide quantitative information about nuclear morphology and pleomorphism, adding histological insights to radiological interpretation of UPS.

Pathologists' perspective on the study design, analysis, and interpretation of proliferative lesions in lifetime and prenatal rodent carcinogenicity bioassays of aspartame.

Aspartame, an artificial sweetener commonly used as a sugar substitute, is currently authorized for use in more than 100 countries. Hundreds of studies, conducted in various countries dating back to the 1970s, have shown that aspartame is safe at real-world exposure levels. Furthermore, multiple human epidemiology studies have provided no indication that consumption of aspartame induces cancer. Given the continued controversy surrounding the Ramazzini Institute's (RI) studies suggesting that aspartame is a carcinogenic hazard in rodents and evaluation by the International Agency for Research on Cancer, this report aims to provide the perspective of experienced pathologists on publicly available pathology data regarding purported proliferative lesions in liver, lung, lymphoid organs, and mammary gland as well as their implications for human risk assessment as reported for three lifetime rodent carcinogenicity bioassays of aspartame conducted at the RI. In the authors' view, flaws in the design, methodology and reporting of the RI aspartame studies limit the utility of the data sets as evidence that this agent represents a carcinogenic hazard. Therefore, all three RI studies, and particularly the accuracy of their pathology diagnoses and interpretations, should be rigorously reviewed by qualified and experienced veterinary toxicologic pathologists in assessing aspartame's carcinogenic risk.